During the next year it is planned to make purified preparations of the alpha, Beta, and gamma subunits. Each of these will be further characterized physically with particular regard to the number of reactive sulfhydryls, the alpha-helical content, as measured by circular dichroism; the internal flexibility, as monitored by fluorescence anisotropy decay; the shape parameters, as measured by the frictional ratio; and the emmission characteristics of the tryptophan groups. In addition confirmatory evidence will be sought for the contention by Graves (JBC, in press) that the catalytic site is on the gamma subunit; this conflicts with an earlier report that it is located on the Beta subunit. It is also planned to attempt to recombine the alpha, Beta and gamma subunits in order to see whether a native kinase molecule with the original activation properties can be reassembled. If this can be done, then assembly will be made with a fluorescent probe located specifically on each of the subunits. In this way the acquisition of rotational mobility by each particular subunit upon activation will be tested for by fluorescence anistropy decay.